This proposal investigates the molecular mechanisms that regulate biological clocks, from the perception of environmental inputs and cellular signals to the initiation of downstream gene expression changes. Biological clocks function in the coordination of gene expression, physiological, and behavioral events at critical times of the day, contributing to adaptive advantages by allowing for the anticipation of daily environmental changes. Substantial progress in the investigation of biological clocks has identified a transcriptional feedback loop, a mechanism conserved throughout numerous organisms. However, less is known about how external stimuli, such as light, are transmitted to the clock, as well as what signal transduction pathways are responsible for transmitting these stimuli. Furthermore, little is known about the functions of the genes whose expression is controlled by the clock. In Drosophila, we have shown that dCLOCK and dCYCLE activate transcription together by binding to E-boxes in the promoters of period and timeless in in vitro expression assays. With this in vitro assay, we have also identified the PER/TIM complex as a potent inhibitor of dCLK/dCYC and determined that the photoreceptor, cryptochrome (CRY), can both bind to TIM and relieve PER/TIM inhibition in light-dependent manners. We have also implicated CBP/p300, a component of the CREB-signaling pathway, in regulating the activity of dCLK/dCYC. Finally, we have used high density oligonucleotide arrays to identify circadianly-expressed genes under clock control. In addition, we propose to investigate the molecular functions of two clock-controlled genes identified in our genome-wide analysis. Our analysis will provide substantial insight into the molecular regulation of circadian clocks and novel information on the links between phototransduction and the regulation of gene expression. Furthermore, because many of these components, pathways, and mechanisms are conserved, our investigation will have significant impact on the understanding of biological clocks and the human diseases that occur from circadian dysfunction.